Summary Motoneuron differentiation and the formation of specific synaptic contacts arise from interactions between expression of genetic information, and epigenetic influences that are extrinsic to the neuron. The objectives of this proposal are to investigate and determine the mechanisms underlying neuronal differentiation and synaptogenesis in mammalian spinal cords. Studies are focused on the mechanisms by which newly formed central and peripheral synapses regulate the developmental changes in motoneuron electrical and pharmacological properties. Development-related increases in motoneuron excitability and in the efficacy of synaptic transmission will be studied using electrophysiological recording techniques. The importance of intracellular Ca2+ in increasing the efficacy of developing sensory-motor synapses will be determined by relating the developmental changes in spontaneous and pharmacologically induced changes in intracellular Ca2+ to physiological changes in synaptic transmission. Three preparations will be used: (l) isolated lumbar spinal cords with attached muscle nerves, (2) spinal cord slices in which sensory-motor pathways are preserved and neurons can be visualized for electrophysiological recordings and Ca2+ imaging, and (3) spinal cord explants in which motoneurons and synaptic pathways differentiate in the controlled environment of organ culture. Our recent studies established the pattern and time course of motoneuron development and synapse formation in utero, and will guide our studies on motoneuron development in organ culture. The proposed experiments are designed to increase our understanding of the roles of extracellular factors in synaptogenesis and neuronal differentiation. This knowledge will provide insight into mechanisms underlying long-term synaptic interactions between neurons in the central nervous system.